Background. More than 800 mutations of the cystic fibrosis gene have been discovered until now. The frequency of these mutations varies widely among cystic fibrosis chromosome of patients from different geographical areas. Both the large number and the heterogeneity of distribution of these mutations require tailoring of specific diagnostic assays. Methods. A diagnostic assay has been designed to allow the simultaneous detection of the 15 most frequent mutations in North-eastern Italy by applying the reverse dot-blot (RDB) hybridization technique after co-amplification in a single tube of 11 PCR products. The RDB assay has been rendered suitable for the analysis of purified DNA and dried blood spots punched from filter paper (Guthrie cards). Results. The RDB assay applied to routine diagnostic testing in 403 cystic fibrosis patients allowed the detection of mutations in 82% of the chromosomes. In a sample of the general population, the assay identified 15 healthy carriers over 443 subjects (1/29.5). Finally, 24 cystic fibrosis patients and 33 healthy carriers were found among 459 newborn babies who have been screened because of elevated blood immunoreactive trypsinogen concentration. Conclusions: The RDB assay proposed for cystic fibrosis mutation detection in our area offers advantages in respect to both detection rate and costs in comparison to the commercially available diagnostic kits. The application of the RDB technique reported here for cystic fibrosis gene suggests the advantages of this relatively inexpensive technology when flexibility and rapid application of research findings to diagnostic protocols is crucial.

Background. More than 800 mutations of the cystic fibrosis gene have been discovered until now. The frequency of these mutations varies widely among cystic fibrosis chromosome of patients from different geographical areas. Both the large number and the heterogeneity of distribution of these mutations require tailoring of specific diagnostic assays. Methods. A diagnostic assay has been designed to allow the simultaneous detection of the 15 most frequent mutations in North-eastern Italy by applying the reverse dot-blot (RDB) hybridization technique after co-amplification in a single tube of 11 PCR products. The RDB assay has been rendered suitable for the analysis of purified DNA and dried blood spots punched from filter paper (Guthrie cards). Results. The RDB assay applied to routine diagnostic testing in 403 cystic fibrosis patients allowed the detection of mutations in 82% of the chromosomes. In a sample of the general population, the assay identified 15 healthy carriers over 443 subjects (1/29.5). Finally, 24 cystic fibrosis patients and 33 healthy carriers were found among 459 newborn babies who have been screened because of elevated blood immunoreactive trypsinogen concentration. Conclusions: The RDB assay proposed for cystic fibrosis mutation detection in our area offers advantages in respect to both detection rate and costs in comparison to the commercially available diagnostic kits. The application of the RDB technique reported here for cystic fibrosis gene suggests the advantages of this relatively inexpensive technology when flexibility and rapid application of research findings to diagnostic protocols is crucial.